UNASSIGNED:CD55在结肠癌的发生发展中起着重要作用。本研究旨在评估CD55在结肠癌中的表达,并发现其如何受到转录因子和miRNA的调控。
未经批准:通过TIMER2.0,UALCAN,和人类蛋白质图谱(HPA)数据库。TRANSFAC和Contrav3用于预测CD55启动子中转录因子的潜在结合位点。TargetScan和starBasev2.0用于预测miRNA与CD55的3'非翻译区(3'UTR)的潜在结合能力。SurvivalMeth用于探索CD55启动子中的差异甲基化位点。Western印迹法检测TFCP2和CD55的表达。双荧光素酶报告基因测定和染色质免疫沉淀(ChIP)测定以确定TFCP2、NF-κB的靶向关系,或miR-27a-3p与CD55。通过构建蛋白质-蛋白质相互作用(PPI)网络并通过基因本体论(GO)和京都基因和基因组百科全书(KEGG)进行途径分析来探索CD55相关基因。
UNASSIGNED:CD55在结肠癌组织中高表达。si-TFCP2降低了TFCP2的mRNA和蛋白表达水平。NF-κB抑制剂明显降低NF-κBmRNA,NF-κB激活剂升高。CD55蛋白也被miR-27a-3p抑制。双荧光素酶报告基因检测表明,敲低TFCP2或抑制NF-κB后,CD55的启动子活性分别下降了21%和70%,分别激活NF-κB后,CD55的启动子活性增加了2.3倍。当TFCP2或NF-κB结合位点突变时,CD55的转录活性明显降低。ChIP实验表明TFCP2和NF-κB结合到CD55的启动子上。与miR-27a-3p模拟物共转染后,CD553'UTR的荧光素酶活性降低,而miR-27a-3pantagomir则增加。随着miR-27a-3p结合位点的突变,我们未发现miR-27a-3p对报道分子活性有显著影响.PPI网络检测揭示了一组CD55相关基因,其中包括CFP,CFB,C4A,C4BGO和KEGG分析显示,靶基因在免疫相关途径中更频繁地出现。
UNASSIGNED:我们的结果表明CD55受TFCP2,NF-κB的调节,miR-27a-3p,和几个免疫相关的基因,进而影响结肠癌。
CD55 plays an important role in the development of colon cancer. This study aims to evaluate the expression of CD55 in colon cancer and discover how it is regulated by transcriptional factors and miRNA.
The expression of CD55 was explored by TIMER2.0, UALCAN, and Human Protein Atlas (HPA) databases. TRANSFAC and Contra v3 were used to predict the potential binding sites of transcription factors in the CD55 promoter. TargetScan and starBase v2.0 were used to predict the potential binding ability of miRNAs to the 3\' untranslated region (3\'UTR) of CD55. SurvivalMeth was used to explore the differentially methylated sites in the CD55 promoter. Western blotting was used to detect the expression of TFCP2 and CD55. Dual-luciferase reporter assay and chromatin immunoprecipitation (ChIP) assay were performed to determine the targeting relationship of TFCP2, NF-κB, or miR-27a-3p with CD55. CD55-related genes were explored by constructing a protein-protein interaction (PPI) network and performing pathway analysis by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG).
CD55 was highly expressed in colon cancer tissues. The mRNA and protein expression levels of TFCP2 were reduced by si-TFCP2. NF-κB mRNA was obviously reduced by NF-κB inhibitor and increased by NF-κB activator. CD55 protein was also inhibited by miR-27a-3p. Dual-luciferase reporter assays showed that after knocking down TFCP2 or inhibiting NF-κB, the promoter activity of CD55 was decreased by 21% and 70%, respectively; after activating NF-κB, the promoter activity of CD55 increased by 2.3 times. As TFCP2 or NF-κB binding site was mutated, the transcriptional activity of CD55 was significantly decreased. ChIP assay showed that TFCP2 and NF-κB combined to the promoter of CD55. The luciferase activity of CD55 3\'UTR decreased after being co-transfected with miR-27a-3p mimics and increased by miR-27a-3p antagomir. As the miR-27a-3p binding site was mutated, we did not find any significant effect of miR-27a-3p on reporter activity. PPI network assay revealed a set of CD55-related genes, which included CFP, CFB, C4A, and C4B. GO and KEGG analyses revealed that the target genes occur more frequently in immune-related pathways.
Our results indicated that CD55 is regulated by TFCP2, NF-κB, miR-27a-3p, and several immune-related genes, which in turn affects colon cancer.